The long term objective of this research is to characterize axolemmnal and myelin molecules which cause Schwann cells (SC) to proliferate and to understand the mechanism by which these signals are transduced across the Schwann cell plasma membrane. Axolemma- enriched fractions (AEF) from myelinated, non-myelinated and unmyelinated sources will be incubated with cultured SC. After stimulation of SC with AEF, fluorescent dyes will be used to study calcium mobilization and activation of the Na+/H+ antiporter; the pattern of cytoplasmic and membrane phosphorylation by 32P will also be examined. Using fluorescent labelled AEF, we will quantitate the amount of AEF per SC to test the hypothesis that a threshold amount of AEF-SC interaction is required to "trigger" mitosis. The myelin-related mitogens will be solubilized and purified by high performance liquid chromatography (HPLC) followed by biochemical characterization. The role of macrophage versus SC processing of exogenous myelin membrane to produce a SC mitogen will be evaluated via the development of macrophage-free cultures. A transformed SC line will be produced utilizing a new DNA construct containing a metallothionein promoter linked to the large T antigen of SV-40 virus in the absence of divalent cation the SC line should undergo normal differentiation when presented with a myelin-competent neurite. Soluble neuronrelated SC mitogen will be produced from cultured neurons by severing them from their neurites and inhibiting the synthesis of a class of molecules to which they are normally bound. The nature of axonal molecules responsible for three molecular manifestations of SC differentiation will be examined; secretion of Type IV procollogen, aggregation of SC surface laminin into linear arrays and possible increase of SC laminin receptors and increase of myelin specific lipids (galactocerebroside) as visualized immunologically and protein (Po, the major myelin glycoprotein) as determined by increased levels of Po mRNA or increased translation of Po mRNA. It is anticipated that these studies will profice a baseline from which to evaluate how glial cells normally respond to neuronal signals os that we can assess how such signalling may be compromised in cases of uncontrolled SC proliferation as in neurofibromatosis. In addition, this research will clarify the relative roles of macrophages verus SC in the proliferative response which follows the Wallerian degeneration characteristic of peripheral nerve injury.